Background: Volumetric absorptive microsampling (VAMS) is a novel sampling technique for the collection of fixed-volume capillary blood. In this study, a new analytical method was developed and used to quantify 14 different antiepileptic drugs (AEDs) and 2 active metabolites in samples collected by VAMS. These data were compared with concentration measurements in plasma. Methods: The authors developed a selective and sensitive liquid chromatography–mass spectrometry (LC-MS/MS) assay to measure the concentrations of several AEDs in whole blood collected by VAMS, which were compared with a commercially available LC-MS/MS kit for AED monitoring in plasma. Drugs and internal standards were extracted from whole blood/plasma samples by a simple protein precipitation. Results: An LC-MS/MS method analyzing VAMS samples was successfully developed and validated for the determination of various AED concentrations in whole blood according to EMA guidelines for bioanalytical method validation. Extraction recovery was between 91% and 110%. No matrix effect was found. The method was linear for all drugs with R2 ≥0.989 in all cases. Intra-assay and inter-assay reproducibility analyses demonstrated accuracy and precision within acceptance criteria. Carry over and interferences were negligible. No volumetric HCT% bias was found at 3 different HCT values (35%–55%) with recovery being consistently above 87%. Samples are very stable at temperatures ranging from −20°C to 37°C and for a 4-month period. Leftover EDTA samples from 133 patients were tested to determine concentration differences between plasma and whole blood sampled by VAMS. The resulting difference varied less than 15% apart from those drugs with a blood/plasma ratio (R) different from 1. Conclusions: The assay allows for highly sensitive and selective quantification of several AEDs in whole blood samples collected by VAMS. The developed method is accurate and precise and free from matrix effects and volumetric HCT% bias.

Background: Therapeutic drug monitoring (TDM) of antiepileptic drugs (AEDs) is commonly performed on plasma or serum. The use of dried plasma spots (DPSs) could represent a useful tool to facilitate sample shipment to reference laboratories. In this article, the authors describe the application of a commercially available UHPLC-MS/MS method for the determination of 9 commonly prescribed AEDs (levetiracetam, lacosamide, topiramate, ethosuximide, lamotrigine, rufinamide, zonisamide, primidone, and oxcarbazepine and its active metabolite 10-OH-monohydroxycarbazepine) to DPS collected on dried sample spot devices (DSSDs). Method: Fifty microliters of plasma were spotted on DSSD. After being air-dried at room temperature, they were extracted using an organic extraction solution containing the appropriate deuterated internal standards. The chromatographic separation was performed on a UHPLC reversed-phase C-18 column, and the analytes were quantified using a triple quadrupole mass spectrometer (LC-MS/MS). Results: The assay was linear over the concentration ranges tested with a total runtime of 10.3 minutes. Recovery ranged from 93.7% to 106.8%. Intraday and interday precision for all quality control levels, including lower limit of quantification, ranged from 2.1% to 18.4% and 2.1% to 13.2%. Intraday and interday accuracy biases ranged from −11.7% to 14.3% and −9.2% to 8.0%. The absence of matrix effects was also tested and confirmed. Real samples derived from patients under therapy were also analyzed, and the comparison of results obtained from DSSD with those obtained from plasma showed that the 2 matrices were interchangeable. Stability tests performed on both quality controls, and real samples demonstrated that DSSDs can be easily stored and shipped at room temperature for 15 days. Conclusions: The application of the LC-MS/MS method allowed the authors to obtain a very specific, sensitive, and rapid (total runtime = 10.3 minutes) quantification of 9 AEDs starting from very low volumes of plasma samples. The main advantage of DPS over wet samples is room temperature storage and shipment, which lowers shipment costs and makes it suitable for routine TDM. Moreover, in comparison with other alternative matrices, DPS allows for the use of the same therapeutic ranges on which routine TDM is based. DPS on DSSD can thus be considered as a useful and cheap tool for the broader application of TDM.

Glioblastoma is characterized by a high proliferative rate and drug resistance. The standard of care includes maximal safe surgery, followed by radiotherapy and temozolomide chemotherapy. The expression of glutamate receptors has been previously reported in human glioma cell lines. The aim of this study was to examine the cellular effects of perampanel, a broad-spectrum antiepileptic drug acting as an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPA) glutamate receptor antagonist, alone or in combination with temozolomide. Four human glioma cell lines were exposed to different concentrations of perampanel and temozolomide, alone or in combination. The type of drug interaction was assessed using the Chou-Talalay method. Apoptosis, cell cycle perturbation, and glutamate receptors (GluRs) subunit expression were assessed by flow cytometry. Perampanel significantly inhibited the growth, inducing high levels of apoptosis. A strong synergistic effect of the combination of perampanel with temozolomide was detected in U87 and A172, but not in U138. Treatment with perampanel resulted in an increased GluR2/3 subunit expression in U87 and U138. Perampanel displays a pro-apoptotic effect on human glioblastoma cell lines when used alone, possibly due to increased GluR2/3 expression. The observed synergistic effect of the combination of temozolomide with perampanel suggests further investigation on the impact of this combination on oncologic outcomes in glioblastoma.

The assay allows for highly sensitive and selective quantification of perampanel and concomitant AEDs in patient plasma samples and can be easily implemented in clinical settings. Our findings are in agreement with previously published data in patients comedicated with enzyme inducer AEDs, but seem to indicate a possible interaction in patients treated with the enzyme inhibitor drug valproic acid.

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